Abstract Apolipoprotein (apo) E is a lipid transport protein that is considered a longevity factor. While the detrimental effects of the human apoE4 isoform have been recognized and studied for quite some time, only recently has apoE2 emerged as being associated with healthy aging and exceptional old age. The proposed studies seek to identify underlying cellular mechanisms by which apoE2 exerts its beneficial effects. Study aims will be advanced using cell- and mouse-models expressing each of the human apoE isoforms. The overarching hypothesis of these studies is that cellular apoE isoforms modulate the expression and activity of membrane ABC transporters, with apoE2 showing prominent positive effects. These membrane proteins mediate the transport of solutes across cellular membranes, and therefore play central roles in biology, maintaining healthy aging and protecting against numerous diseases such as diabetes, cardiovascular and Alzheimer's diseases. As such, an understanding of how cellular apoE regulates ABC transporters will facilitate the identification of new therapeutic- and diagnostic- targets to combat disease. A series of cell-based, hypothesis-driven studies are proposed to investigate the effects of each apoE isoform on specific ABC transporters involved in lipid homeostasis. Targeted ?omic? technologies will be used to define metabolic phenotypes associated with each apoE isoform, which tract to ABC transporter function. Novel cryo-EM and 3D modeling techniques will be employed to investigate the ultrastructure of cells, and whether the size, shape, and number of cytoplasmic organelles correlate with these metabolic phenotypes. Experiments will be performed under basal and oxidative stress conditions, thereby providing detailed insights into the function of cells, their resilience, and the changes that occur during disease development in relation to each of the human apoE isoforms. Mimetic peptides based on favorable apoE2 responses will be developed, and used to investigate structure-activity relationships. In vivo mouse studies will also be performed to validate new mechanisms associated with apoE2/mimetic peptides, identifying metabolite- and lipid-biomarkers linked to the function of specific ABC transporters for clinical applications.